Atomic Structure of the Murine Norovirus Protruding Domain and Soluble CD300lf Receptor Complex

Abstract

Human noroviruses are the leading cause of acute gastroenteritis in humans. Noroviruses also infect animals, such as cows, mice, cats, and dogs. How noroviruses bind and enter host cells is still incompletely understood. Recently, the type I transmembrane protein CD300lf was identified as the murine norovirus receptor, yet it is unclear how the virus capsid and receptor interact at the molecular level. In this study, we determined the X-ray crystal structure of the soluble CD300lf (sCD300lf) and the murine norovirus capsid protruding domain complex at a 2.05-Å resolution. We found that the sCD300lf-binding site is located on the topside of the protruding domain and involves a network of hydrophilic and hydrophobic interactions. sCD300lf locked nicely into a complementary cavity on the protruding domain that is additionally coordinated with a positive surface charge on sCD300lf and a negative surface charge on the protruding domain. Five of six protruding domain residues interacting with sCD300lf were maintained between different murine norovirus strains, suggesting that sCD300lf was capable of binding to a highly conserved pocket. Moreover, a sequence alignment with other CD300 paralogs showed that the sCD300lf-interacting residues were partially conserved in CD300ld but variable in other CD300 family members, consistent with previously reported infection selectivity. Overall, these data provide insights into how a norovirus engages a protein receptor and will be important for a better understanding of selective recognition and norovirus attachment and entry mechanisms. Noroviruses exhibit exquisite host range specificity due to species-specific interactions between the norovirus capsid protein and host molecules. Given this strict host range restriction, it has been unclear how the viruses are maintained within a species between relatively sporadic epidemics. While much data demonstrate that noroviruses can interact with carbohydrates, recent work has shown that expression of the protein CD300lf is both necessary and sufficient for murine norovirus infection of mice and binding of the virus to permissive cells. Importantly, the expression of this murine protein by human cells renders them fully permissive for murine norovirus infection, indicating that at least in this case, host range restriction is determined by molecular events that control receptor binding and entry. Defining the atomic-resolution interactions between the norovirus capsid protein and its cognate receptor is essential for a molecular understanding of host-range restriction and norovirus tropism.

High-resolution cryo-EM structures of outbreak strain human norovirus shells reveal size variations

“…Recently reported co-crystal structures of murine norovirus P-domains in complex with their protein receptor, CD300lf, revealed the receptors bind at the interfaces between three P-domain spikes when superimposed on the map of the assembled virus (19)(20)(21). In GII.4 T=4 particles, the interface between dimeric Pdomain spikes forms around the 20 icosahedral three-fold axes between Dsubunits, and the 60 quasi three-fold axes between A, B and C subunits (Fig.…”

“…The addition of either EDTA or 1,10-phenanthroline that preferentially chelates Zn 2+ caused an over three-fold decrease in the number of intact VLPs observed and the metal density disappeared in the asymmetric and symmetric reconstructions from the remaining intact particles, whereas EGTA, which preferentially chelates Ca 2+ , did not cause such changes. The infectivity of murine norovirus and feline caliciviruses that do not bind HBGAs have been reported to be metal-dependent (19,20,22). Moreover, GII.2 VLPs were reported to bind HBGAs when mixed with GII.2-positive patient stool, underscoring the involvement of co-ligands (23).…”

CD300lf is the primary physiologic receptor of murine norovirus but not human norovirus

“…P2 is the target for several neutralizing monoclonal antibodies for both HNoV and MNoV and contains the receptor binding site for MNoV [12,13]. Co-crystal structures and mutagenesis studies have identified that the CC' and CDR3 loops of CD300lf directly bind to a cleft between the AB' and DE' loops of the MNoV VP1 P2 subdomain [13,15,19]. Each CD300lf binds one P2 monomer, albeit at relatively low affinity, suggesting that norovirus-receptor interactions are largely driven by avidity [13].…”

Heterologous Expression of Human Norovirus GII.4 VP1 Leads to Assembly of T=4 Virus-Like Particles

“…Structural studies have shown that caliciviruses have a common overall organization of T=3 icosahedral symmetry and are comprised of 180 copies of VP1 (7, 8, 23-25). Within the asymmetric unit, VP1 adopts three quasi-equivalent conformations, termed A, B, and C (8).…”